Our laboratory has a long-standing interest in immune tolerance, with a particular focus on mechanisms that maintain self-tolerance. Multiple receptors which regulate T cell development and mature T cell responses (e.g., TCR, CD28, and &#947;c cytokine receptors) signal via PI-3 kinase (PI3K). The tumor suppressor gene Pten is a lipid phosphatase which directly catalyzes the reverse of the PI3K reaction. Previously, we determined that PTEN is expressed in na[unreadable]ve and regulatory T cells. Using mice with PTEN deletion targeted to the T cell compartment (PTEN-&#916;T mice), we found that PTEN regulates both TCR and IL-2R mediated responses. In the absence of PTEN, na[unreadable]ve T cells could be optimally activated in vitro and in vivo without CD28 costimulation, and regulatory T cells could be induced to proliferate to IL-2 without a need for TCR activation. Thus PTEN is a critical regulator of mature T cell homeostasis and function. As they age PTEN-&#916;T mice develop T cell lymphomas and a generalized autoimmune syndrome. Previous models suggested that this was due to the lipid phosphatase activity of PTEN, which modulated TCR and growth factor receptor signaling. However new data indicates that PTEN has a nuclear function as well, where it binds to centromeres and mediates chromosomal stability. Interestingly, our preliminary data indicate that lymphoma development is an intrathymic event while autoimmunity arises from mature peripheral T cells, suggesting that distinct mechanisms govern these two consequences of PTEN-deficiency in T cells. Based on these data, we have two aims. In aim #1, we will test the hypothesis that distinct PTEN functions are responsible for prevention of autoimmunity and lymphoma. This will be done using mice in which mutant forms have Pten, lacking phosphatase function or nuclear localization, have been knocked into the endogenous Pten locus. Additional studies will examine the role of TCR signaling in tumor growth and survival. In aim #2, we will use will use a foxp3-YFP-Cre transgenic mouse and Ptenflox mice to delete PTEN in the major subset of Tregs. Introduction of a Rosa26R-GFP reporter will permit fate mapping. We will use these mice to study three areas: the role of PTEN in in vivo nTreg and iTreg development and lineage stability;whether loss of PTEN obviates the need for CD28 signals in Treg development or maintenance;and how loss of PTEN in Tregs influences susceptibility to autoimmunity, using the NOD model. Together, studies in these two aims will provide key new insights into the role of Pten in T cell development, homeostasis, and protection from autoimmunity and lymphoma.